Binding of eukaryotic initiation factor 3 to ribosomal 40S subunits and its role in ribosomal dissociation and anti-association
Binding of eukaryotic initiation factor 3 to ribosomal 40S subunits and its role in ribosomal dissociation and anti-association
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DOI：
10.1261/rna.7215305
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发表时间：
2005-04-01
期刊：
影响因子：
4.5
通讯作者：
Pestova, TV
中科院分区：
文献类型：
--
作者：
Kolupaeva, VG;Unbehaun, A;Pestova, TV
多亚基真核起始因子（eIF） 3在翻译起始过程中发挥多种作用，均涉及与40S核糖体亚基的相互作用。eIF3可以以两种形式纯化：带或不带松散关联的eIF3j亚基（分别为eIF3j+和eIF3j-）。尽管与eIF3j+不同，eIF3j-与40S亚基的结合不够稳定，无法承受糖密度梯度离心作用，但我们发现，除了已知的eIF2-GTP-Met-tRNA(i)（Met）三元配合物对eIF3/40S亚基相互作用的稳定作用外，eIF3j-/40S亚基配合物还可以被至少25 nt长的单链RNA或DNA辅助因子稳定，这些辅助因子可以被稳定的发夹夹在两侧。在所有均聚物中，oligo（rU）、oligo（dT）和oligo（dC）刺激了eIF3/40S亚基相互作用，而oligo（rA）、oligo（rG）、oligo（rC）、oligo（dA）和oligo（dG）则没有。其他碱基穿插的寡核苷酸(U)或寡核苷酸（dT）序列也促进了这种相互作用。寡核苷酸刺激eIF3/40S亚基结合的能力与其与40S亚基结合的能力相关，最有可能与40S亚基的mrna结合间隙有关。虽然eIF3j+可以直接结合40S亚基，但无论是eIF3j-还是eIF3j+都不能分离80S核糖体，也不能保护40S和60S亚基不重新结合。值得注意的是，在eif2三元配合物或任何促进eIF3/ 40s亚基相互作用的寡核苷酸辅因子存在时，两种形式的eIF3的解离/抗缔合活性都变得明显。eIF1显著增强了eIF3的核糖体解离和抗结合力。在缺乏eif2三元配合物的情况下，通过RNA辅助因子刺激eIF3/40S亚基相互作用的潜在生物学作用进行了讨论。
The multisubunit eukaryotic initiation factor (eIF) 3 plays various roles in translation initiation that all involve interaction with 40S ribosomal subunits. eIF3 can be purified in two forms: with or without the loosely associated eIF3j subunit (eIF3j+ and eIF3j-, respectively). Although unlike eIF3j+, eIF3j- does not bind 40S subunits stably enough to withstand sucrose density gradient centrifugation, we found that in addition to the known stabilization of the eIF3/40S subunit interaction by the eIF2-GTP-Met-tRNA(i)(Met) ternary complex, eIF3j-/40S subunit complexes were also stabilized by single-stranded RNA or DNA cofactors that were at least 25 nt long and could be flanked by stable hairpins. Of all homopolymers, oligo(rU), oligo(dT), and oligo(dC) stimulated the eIF3/40S subunit interaction, whereas oligo(rA), oligo(rG), oligo(rC), oligo(dA), and oligo(dG) did not. Oligo(U) or oligo(dT) sequences interspersed by other bases also promoted this interaction. The ability of oligonucleotides to stimulate eIF3/40S subunit association correlated with their ability to bind to the 40S subunit, most likely to its mRNA-binding cleft. Although eIF3j+ could bind directly to 40S subunits, neither eIF3j- nor eIF3j+ alone was able to dissociate 80S ribosomes or protect 40S and 60S subunits from reassociation. Significantly, the dissociation/anti-association activities of both forms of eIF3 became apparent in the presence of either eIF2-ternary complexes or any oligonucleotide cofactor that promoted eIF3/40S subunit interaction. Ribosomal dissociation and anti-association activities of eIF3 were strongly enhanced by eIF1. The potential biological role of stimulation of eIF3/40S subunit interaction by an RNA cofactor in the absence of eIF2-ternary complex is discussed.